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Role of transient receptor potential vanilloid 1 in the modulation of airway smooth muscle tone and calcium handling.

ABSTRACT: Asthma is a common disorder characterized, in part, by airway smooth muscle (ASM) hyperresponsiveness. Transient receptor potential vanilloid 1 (TRPV1) is a nonselective cation channel expressed on airway nerve fibers that modulates afferent signals, resulting in cough, and potentially bronchoconstriction. In the present study, the TRPV1 transcript was detected by RT-PCR in primary cultured human ASM cells, and the TRPV1 protein was detected in ASM of human trachea by immunohistochemistry. Proximity ligation assays suggest that TRPV1 is expressed in the sarcoplasmic reticulum membrane of human ASM cells in close association with sarco/endoplasmic reticulum Ca2+-ATPase-2. In guinea pig tracheal ring organ bath experiments, the TRPV1 agonist capsaicin led to ASM contraction, but this contraction was significantly attenuated by the sodium channel inhibitor bupivacaine (n = 4, P < 0.05) and the neurokinin-2 receptor antagonist GR-159897 (n = 4, P < 0.05), suggesting that this contraction is neutrally mediated. However, pretreatment of guinea pig and human ASM in organ bath experiments with the TRPV1 antagonist capsazepine inhibited the maintenance phase of an acetylcholine-induced contraction (n = 4, P < 0.01 for both species). Similarly, capsazepine inhibited methacholine-induced contraction of peripheral airways in mouse precision-cut lung slice (PCLS) experiments (n = 4-5, P < 0.05). Although capsazepine did not inhibit store-operated calcium entry in mouse ASM cells in PCLS (n = 4-7, P = nonsignificant), it did inhibit calcium oscillations (n = 3, P < 0.001). These studies suggest that TRPV1 is expressed on ASM, including the SR, but that ASM TRPV1 activation does not play a significant role in initiation of ASM contraction. However, capsazepine does inhibit maintenance of contraction, likely by inhibiting calcium oscillations.

SUBMITTER: Yocum GT

PROVIDER: S-EPMC5495950 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Role of transient receptor potential vanilloid 1 in the modulation of airway smooth muscle tone and calcium handling.

Yocum Gene T GT Chen Jun J Choi Christine H CH Townsend Elizabeth A EA Zhang Yi Y Xu Dingbang D Fu Xiao W XW Sanderson Michael J MJ Emala Charles W CW

American journal of physiology. Lung cellular and molecular physiology 20170323 6

Asthma is a common disorder characterized, in part, by airway smooth muscle (ASM) hyperresponsiveness. Transient receptor potential vanilloid 1 (TRPV1) is a nonselective cation channel expressed on airway nerve fibers that modulates afferent signals, resulting in cough, and potentially bronchoconstriction. In the present study, the TRPV1 transcript was detected by RT-PCR in primary cultured human ASM cells, and the TRPV1 protein was detected in ASM of human trachea by immunohistochemistry. Proxi ...[more]

PMID: 28336810

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Project description:ObjectvesTransient receptor potential vanilloid 3 (TRPV3) is a member of the TRP channels family of Ca2+ -permeant cation channels. In this study, we aim to investigate the role of TRPV3 in pulmonary vascular remodeling and PASMCs proliferation under hypoxia.Materials and methodsThe expression of TRPV3 was evaluated in patients with pulmonary arterial hypertension (PAH) and hypoxic rats, using hematoxylin and eosin (H&E) and immunohistochemistry. In vitro, MTT assay, flow cytometry, Western blotting and immunofluorescence were performed to investigate the effects of TRPV3 on proliferation of PASMCs.ResultsWe found that, in vivo, the expression of TRPV3 was increased in patients with PAH and hypoxic rats. Right ventricular hypertrophy measurements and pulmonary pathomorphology data show that the ratio of the heart weight/tibia length (HW/TL), the right ventricle/left ventricle plus septum (RV/LV+S) and the medial width of the pulmonary artery were increased in chronic hypoxic rats. Moreover, the expression of proliferating cell nuclear antigen (PCNA), Cyclin D, Cyclin E and Cyclin A, phospho-CaMKII (p-CaMKII) were induced by hypoxia. In vitro, we revealed that hypoxia promoted PASMCs viability, increased the expression of PCNA, Cyclin D, Cyclin E, Cyclin A p-CaMKII, made more cells from G0 /G1 phase to G2 /M + S phase, enhanced the microtubule formation, and increased [Ca2+ ]i , which could be suppressed by Ruthenium Red, an inhibitor of TRPV3, and TRPV3 silencing has similar effects. Furthermore, the up-regulated expression of PCNA, Cyclin D, Cyclin E and Cyclin A, the increased number of cells in G2 /M and S phase, and the enhanced activation and expression of PI3K and AKT proteins induced by hypoxia and in presence of carvacrol (an agonist of TRPV3), was significantly attenuated by incubation of LY 294002, a specific inhibitor for PI3K/AKT.ConclusionsThese findings suggest that TRPV3 is involved in hypoxia-induced pulmonary vascular remodeling and promotes proliferation of PASMCs and the effect is, at least in part, mediated via the PI3K/AKT pathway.

Project description:Chronic hypoxia causes remodeling and alters contractile responses in both pulmonary arteries and pulmonary veins. Although pulmonary arteries have been studied extensively in these disorders, the mechanisms by which pulmonary veins respond to hypoxia and whether these responses contribute to chronic hypoxic pulmonary hypertension remain poorly understood. In pulmonary arterial smooth muscle, we have previously demonstrated that influx of Ca(2+) through store-operated calcium channels (SOCC) thought to be composed of transient receptor potential (TRP) proteins is likely to play an important role in development of chronic hypoxic pulmonary hypertension. To determine whether this mechanism could also be operative in pulmonary venous smooth muscle, we measured intracellular Ca(2+) concentration ([Ca(2+)](i)) by fura-2 fluorescence microscopy in primary cultures of pulmonary venous smooth muscle cells (PVSMC) isolated from rat distal pulmonary veins. In cells perfused with Ca(2+)-free media containing cyclopiazonic acid (10 ?M) and nifedipine (5 ?M) to deplete sarcoplasmic reticulum Ca(2+) stores and block voltage-dependent Ca(2+) channels, restoration of extracellular Ca(2+) (2.5 mM) caused marked increases in [Ca(2+)](i), whereas MnCl(2) (200 ?M) quenched fura-2 fluorescence, indicating store-operated Ca(2+) entry (SOCE). SKF-96365 and NiCl(2), antagonists of SOCC, blocked SOCE at concentrations that did not alter Ca(2+) responses to 60 mM KCl. Of the seven known canonical TRP (TRPC1-7) and six vanilloid-related TRP channels (TRPV1-6), real-time PCR revealed mRNA expression of TRPC1 > TRPC6 > TRPC4 > TRPC2 ? TRPC5 > TRPC3, TRPV2 > TRPV4 > TRPV1 in distal PVSMC, and TRPC1 > TRPC6 > TRPC3 > TRPC4 ? TRPC5, TRPV2 ? TRPV4 > TRPV1 in rat distal pulmonary vein (PV) smooth muscle. Western blotting confirmed protein expression of TRPC1, TRPC6, TRPV2, and TRPV4 in both PVSMC and PV. Our results suggest that SOCE through Ca(2+) channels composed of TRP proteins may contribute to Ca(2+) signaling in rat distal PV smooth muscle.

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Role of transient receptor potential vanilloid 1 in the modulation of airway smooth muscle tone and calcium handling.

Publications

Role of transient receptor potential vanilloid 1 in the modulation of airway smooth muscle tone and calcium handling.

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